Carcinogenesis is a multistage event affected by a variety of genetic and epigenetic factors and is typified by the outbreak of uncontrolled cell growth originated from different tissues. A universal goal for anticancer research lies in the development of a clinical treatment that is highly effective in curtailment of tumor growth, non-toxic to the host, and is affordable for most patients. Drugs that inhibit targets that are unique to dividing cells, particularly cells dividing in an uncontrolled manner, are an ideal paradigm for chemotherapeutic agents, the greater the specificity to cells that are dividing in an uncontrolled manner the lower the risk of attendant side effects.
The inventors and colleagues have previously reported that the tetra-O-methyl nordihydroguaiaretic acid (M4N), also known as EM1421 and terameprocol, a semi-chemically synthesized derivative of nordihydroguaiaretic acid (NDGA) possessed antiviral (1, 2) and anti-cancer (3) activities in cultured cells, in mouse models (3, 4), and in human xenografts in nude mice (5). As a transcription inhibitor, M4N suppresses Sp1-regulated cdk expression and causes cell cycle arrest at the G2 phase of the cell cycle (3, 4). The overexpression of Sp1 has been shown to have significant roles in development and progression of gastric cancer (6, 7). The safety and possible tumoricidal capability of M4N was examined for human patients either through intralesional (8) or topical applications (9). M4N currently undergoes Phase I/II clinical trials in patients by intravenous infusion (10). The clinical trial data so far indicated that M4N had substantial anticancer efficacy. However the data also suggested that it is desirable if we can find some ways to strengthen the anticancer activity of the drug since the tumoricidal efficacy of this drug is not strong enough in most of cases. There are many reports to indicate that the combination drug regimens based on several anticancer drugs are effective for the treatment of certain type of cancers (11, 12). In this study we explored possible anticancer combination drug treatments based on M4N, and investigated mechanistic backgrounds for effective anticancer therapy. Inhibiting cell death is widely accepted as a necessary step in the transition from normal to cancer cells, and most cancer therapies exert their effects by indirectly reversing this process (13, 14). Mitochondria are known to be a key element in cell death mechanisms. The activation of Apaf-1, caspase-9 and -3, and ICAD (DFF45) by cytochrome c egress from mitochondria is considered to be the most important mechanism for apoptosis induction (15). Usually mitochondrial membrane potential depolarization accompanies activation of cell death signal in mitochondria (16, 17). In many cancer cells, mitochondrial membrane potential is hyperpolarized rather than depolarized, which is one of many indications implying that cell death mechanism at the mitochondrial level is often broken in cancer (13, 14). For this reason it is quite understandable why many efforts of anticancer drug development are focused on normalizing mitochondria-related cell death pathway (18-20). Bcl-2 family proteins which are either pro-apoptotic or anti-apoptotic regulate cell death at the mitochondrial level (21). BNIP-3 is a protein belonging to Bcl-2 family protein family. This protein is a pro-apoptotic protein which is considered to mediate cell death signal through mitochondria under the stress conditions, most notably under hypoxic condition. Since cancer cells are often exposed to hypoxic condition because tumors often don't get enough blood oxygen supply, the study on this particular protein is very important to understand pathology of cancer development and prevention.
In recent years many cancer researchers have investigated extensively about autophagy as well as cell death, considering the exploitation of both of these physiological mechanisms to be crucial for establishing effective anticancer therapeutic regimens (27, 28). In the literatures there is clear evidence from the phenotypes of mutant mice, and cells derived from the mice, that autophagy functions to sustain cell survival, particularly during stress (29-36). It is also clear that there is functional interaction between autophagy and cell death pathways (37, 38). In response to metabolic stress, autophagy can delay cell death by apoptosis, and in apoptotic-defective cells, inactivation of the autophagy survival pathway promotes necrotic cell death in vitro and in tumors in vivo (29, 31, 38). Considering this recent progress in cancer research, mechanistic perspectives of both cell death and autophagy will be addressed for establishing effective combination drug regimens based on M4N in this study.
This application claims priority to U.S. Provisional Patent Application 61/306,680, filed Feb. 22, 2010, which is incorporated by reference in its entirety.